Many of the current shortcomings in diagnosis, prognosis, risk stratification and treatment of disease can be approached through the identification of the molecular mechanisms underlying a disease and through the discovery of nucleotide sequences (or sets of nucleotide sequences) whose expression patterns predict the occurrence or progression of disease states, or predict a patient's response to a particular therapeutic intervention. In particular, identification of nucleotide sequences and sets of nucleotide sequences with such predictive value from cells and tissues that are readily accessible would be extremely valuable. For example, peripheral blood is attainable from all patients and can easily be obtained at multiple time points at low cost. This is a desirable contrast to most other cell and tissue types, which are less readily accessible, or accessible only through invasive and aversive procedures. In addition, the various cell types present in circulating blood are ideal for expression profiling experiments as the many cell types in the blood specimen can be easily separated if desired prior to analysis of gene expression. While blood provides a very attractive substrate for the study of diseases using expression profiling techniques, and for the development of diagnostic technologies and the identification of therapeutic targets, the value of expression profiling in blood samples rests on the degree to which changes in gene expression in these cell types are associated with a predisposition to, and pathogenesis and progression of a disease.
There is an extensive literature supporting the role of leukocytes, e.g., T- and B-lymphocytes, monocytes and granulocytes, including neutrophils, in a wide range of disease processes, including such broad classes as cardiovascular diseases, inflammatory, autoimmune and rheumatic diseases, infectious diseases, transplant rejection, cancer and malignancy, and endocrine diseases.
Of particular interest is the role of leukocytes and leukocyte gene expression in chronic inflammatory diseases such as Systemic Lupus Erythematosis and Rheumatoid Arthritis. Systemic lupus erythematosis (SLE) and Rheumatoid Arthritis (RA) are chronic autoimmune and inflammatory disorders characterized by dysregulation of the immune system, which causes damage to a variety of organs. These diseases clearly involve differential expression of genes in leukocytes. Diagnostic and disease monitoring tools are severely lacking for these patients and their physicians. Leukocyte expression profiling can be applied to discover expression markers for SLE and RA and apply them as patient management tools in the clinical setting. In addition, osteoarthritis is a degenerative joint disease that can be confused with RA. This disease also involves leukocytes and expression profiling of leukocytes associated with osteoarthritis may lead to the discovery of new diagnostic and therapeutic approaches to the disease.
SLE in particular is a chronic, usually life-long, potentially fatal autoimmune disease characterized by unpredictable exacerbations and remissions with protean clinical manifestations. SLE is notable for unpredictable exacerbations and remissions and a predilection for clinical involvement of the joints, skin, kidney, brain, serosa, lung, heart, and gastrointestinal tract. The pathologic hallmark of the disease is recurrent, widespread, and diverse vascular lesions.
SLE is not a rare disorder. Although reported at both extremes of life (e.g. diagnosed in infants and in the tenth decade of life) chiefly it affects women of child bearing age. Among children, SLE occurs three times more commonly in females than in males. In the 60% of SLE patients who experience onset of their disease between puberty and the fourth decade of life the female to male ratio is 9:1. Thereafter, the female preponderance again falls to that observed in prepubescents.
The disorder is three times more common in African American blacks than American caucasians. SLE is also more common in Asians and in China may be more common than Rheumatoid Arthritis. The ethnic group at greatest risk is African Caribbean blacks. The annual incidence of SLE ranges from six to 35 new cases per 100,000 population in relatively low-risk to high-risk groups. The prevalence of SLE in the United States is an issue of some debate. Prevalence estimates of between 250,000 to 500,000 are contradicted by a recent nationwide telephone poll suggesting a prevalence of between one and two million.
The prognosis for patients with SLE has greatly improved over the last few decades with at least 80-90% of all patients surviving ten years. Thereafter life expectancy approximates that of age matched controls. This improvement reflects the general advancements in health care (i.e. dialysis, antibiotics, antihypertensives, newer immunosuppressives with more favorable efficacy to toxicity ration) but also the specialized care available for patients with SLE.
Such specialized medical care includes care by experienced clinicians with access to state of the art diagnostic and therapeutic measures will result in improved outcomes and the most cost-effective utilization of resources. Expert care of patients with SLE leads to fewer hospitalizations secondary to uncontrolled disease exacerbation, less severe renal disease with fewer patients experiencing end stage renal disease requiring chronic dialysis, fewer episodes of avascular necrosis requiring total joint replacement, and less severe osteoporosis and fractures. In addition, more judicious use of steroids and steroid sparing agents can also reduce the severity of atherosclerosis and resulting incidence of myocardial infarctions and cerebral vascular accidents, which now complicate the natural history of SLE. Improved monitoring, diagnosis and prognosis of SLE should aid clinicians in determining appropriate care for SLE patients, including which drugs to use and at what amounts.
At a molecular level, SLE is an autoimmune disease characterized by immune dysregulation resulting in the production of antinuclear antibodies (ANA), generation of circulating immune complexes, and activation of the complement system. SLE is further characterized by end organ damage that results from deposition of circulating autoantibodies and subsequent complement- and Fc receptor-mediated inflammation. In addition, extensive immune system abnormalities, including altered T lymphocyte function and spontaneous apoptosis, contribute to the lymphopenia and increased susceptibility to infection that confer considerable morbidity.
The clinical features of SLE are protean and may mimic infectious mononucleosis, lymphoma, or other systemic disease. Therefore, the American College of Rheumatology developed criteria to include patients with SLE and exclude those with other disorders. These criteria are best used to insure the appropriateness of subjects for epidemiological or research studies. Although many patients do not fulfill the rigid criteria at first encounter most will when followed over periods of time.
The etiology of SLE remains unknown. A genetic predisposition, sex hormones, and environmental trigger(s) likely result in the disordered immune response that typifies the disease.
A role for genetics is suggested by the increased percentage of two histocompatibility antigens in patients with SLE, HLA-DR2 and HLA-DR3. In addition, there is an increased frequency of the extended haplotype HLA-A1, B8, DR3. The role for heredity is further supported by the concordance for this illness among monozygotic twins. The polygenic nature, however, of this genetic predisposition as well as the contribution of environmental factors is suggested by the only moderate concordance rate which is reported to be between 25 and 60%.
The origin of autoantibody production in SLE is unclear but a role has been suggested for an antigen driven process, spontaneous B-cell hyper-responsiveness, or impaired immune regulation. Regardless of the etiology of autoantibody production, SLE is associated with the impaired clearance of circulating immune complexes secondary to decreased CR1 expression, defective Fc receptor function, or deficiencies of early complement components such as C4A.
More is known about the pathogenic cellular and molecular events that are responsible for vascular lesions in SLE than the origins of autoimmunity. Disease manifestations result from recurrent vascular injury due to immune complex deposition, leukothrombosis, or thrombosis. Additionally, cytotoxic antibodies can mediate autoimmune hemolytic anemia and thrombocytopenia, while antibodies to specific cellular antigens can disrupt cellular function. An example of the latter, is the association between anti-neuronal antibodies and neuropsychiatric SLE.
The health status of a patient with SLE is related not only to disease activity, but to the damage that results from recurrent episodes of disease flare (i.e. deforming arthropathy, shrinking lung, end stage renal disease, organic mental syndrome, etc.), as well as the adverse effects of treatment (i.e. avascular necrosis of bone, infections, and precocious atherosclerosis, etc.).
The accuracy of technologies based on expression profiling for the diagnosis, prognosis, and monitoring of disease would be dramatically increased if numerous differentially expressed nucleotide sequences, each with a measure of sensitivity and specificity for a disease in question, could be identified and assayed in a concerted manner. Using the expression of multiple genes (gene sets) for diagnostic applications helps overcome assay and population variability. PCT application WO 02/057414 “LEUKOCYTE EXPRESSION PROFILING” to Wohlgemuth identifies a set of differentially expressed nucleotides.
In order to achieve this improved accuracy, the appropriate sets of nucleotide sequences once identified need to be validated against numerous samples in combination with relevant clinical data.